This invention relates to electrophoresis processes, and in particular, to a construction for an electrophoresis chamber employed in such processes. While the invention is described with reference to a particular electrophoresis process, those skilled in the art will recognize the wider applicability of the inventive principles disclosed hereinafter.
The free-flow electrophoretic process with which the invention disclosed hereinafter finds application is a combination of several phenomenon. As will be appreciated by those skilled in the art, a free-flow electrophoresis process is a continuous laminar flow of a carrier fluid or buffer that flows through a chamber with multiple outlets. A sample stream containing protein, cells, or other particles forms an input into this flow and an electrical field is applied across the flow in the direction of desired separation. The underlying principle for separation is electrical phoresis, that is to say, the motion of charged particles in an electrical field. The applied field results in a force on the particles, which force is proportional to their charge and the electrical field strength. Under the influence of this force, the particles are rapidly accelerated in the direction of the force approaching their terminal velocity for equilibrium with the viscous drag force on the particles. Different particles will, in general, have different lateral velocities and will exit the chamber through different exit ports, accomplishing separation of the sample. The characteristics used to quantify this phenomenon is particle mobility, which is defined for the purpose of this specification in its normal connotation, as the velocity component in the direction of the electrical field divided by the electrical field strength applied to the particle.
If the carrier buffer is a fluid of constant properties, that is to say, constant density and viscosity, the flow can be characterized as Poiseuille flow or fully developed, one dimensional flow between parallel plates. In actual practice, Poiseuille flow is not obtainable. Departures from Poiseuille flow occur near the inlet to the buffer flow chamber and near the sides of the buffer flow chamber, where the membranes used to isolate the electrodes and the surrounding buffer which contains hydrogen and oxygen gas and other possible breakdown products from the buffer flow in the chamber act as a zero velocity boundary. Because the plate spacing defining the buffer flow chamber is small compared to the chamber width, the maximum fluid velocity at the midpoint between the plates is very nearly constant except near the membranes. If the electrical field strength is constant throughout the chamber, then particles forming an input to the carrier buffer will have constant lateral terminal velocities and their paths will be nearly straight lines running diagonally across the chamber length. Carrier buffer conventionally is mostly water with additives to support the viability of the biological materials being separated. Water definitely is not a fluid of constant properties. For terrestrial application of electrophoresis, the important property variation is the variation of density with temperature. Whenever temperature differences exist at the same altitude within the chamber, the less dense fluid experiences an upward buoyant force due to gravity and the more dense fluid experiences a greater downward buoyant force due to gravity. These gravity forces give rise to convection currents that are superimposed on the main flow. In the past, minimizing convection currents in electrophoresis processes has required either that the temperature differences be minimized or that viscous damping of the currents be increased, or both. Chamber thickness as small as 0.6 millimeter has been used in analytical electrophoresis units to provide added viscous damping of convection currents in the flow. Thin chambers, however, result in sample cross sections too small for practical production of pharmaceutical products.
The invention disclosed hereinafter overcomes these prior art difficulties by providing a chamber that combines several structural features to minimize temperature differences in the direction of separation across the buffer flow chamber. In particular, the chamber of this invention employs a cooling flow that traverses the direction of buffer flow to minimize horizontal temperature differences. It also employs a construction in which the cooling fluid and the buffer fluid surrounding the electrodes outside the membranes are one in the same, eliminating temperature differences in the chamber buffer due to temperature differences between cooling fluid and buffer fluid surrounding the electrodes.
One of the objects of this invention is to provide an improved electrophoresis chamber design.
Another object of this invention is to provide an electrophoresis chamber design in which temperature variations in any particular horizontal plane across the chamber are minimized.
Another object of this invention is to provide an electrophoresis chamber design in which temperature differences due to separate cooling fluid and buffer fluid surrounding the electrodes are eliminated.
Another object of this invention is to provide an electrophoresis chamber design in which a serpentine coolant-buffer flow is provided.
Still another object of this invention is to provide an electrophoresis chamber design employing predeterminedly spaced baffles to channel coolant buffer flow in a desired direction.
Other objects of this invention will be apparent to those skilled in the art in light of the following description and accompanying drawings.